Li-Chien Hsu

Neutrophil dynamics and retention in lung, oxygenator, and arterial filter during cardiopulmonary bypass in a pig model.

Interactions of neutrophils with adsorbed proteins in components of the cardiopulmonary bypass (CPB) circuit and expression of leukocyte adhesion molecules on activated neutrophils affect neutrophil kinetics and margination. Lung and skeletal muscle along with oxygenator (OX) and arterial filter (AF) in the extracorporeal circuit provide the major areas of neutrophil (N) interaction. The dynamics of N-interaction and N-retention during 3 hr CPB was quantified with autologous In-111 labeled neutrophils (INN) in 4 groups of 20 Yorkshire pigs (28-35 kg, 5 sham; 5 CPB, 1 hr; 5 CPB, 3 hr and 5 CPB with heparinized circuit, 3 hr); anesthetized pigs were injected with INN (500-650 microCi), 30 min before CPB and heparinized, and underwent CPB with a roller pump, a hollow fiber OX (Bentley CM 50, 5.0 m2) and AF (Bentley AF 025, 0.25 m2) at 2.5-3.6 l/min for 3 hr. N-dynamics on OX and AF was monitored by a calibrated Geiger probe. Neutrophil deposition, like that of plasma proteins on OX, reached a steady state almost instantly, but increased on filter with CPB time. INN distribution was viewed with a gamma camera; total INN was measured with an ion chamber and INN in samples of fibers and tissues was quantified with a gamma counter. INN in lung did not change significantly during CPB and increased in liver. The percentage of injected INN in lung, liver, and brain changed with CPB time and showed significant increase over sham-operated animals. Heparin coating of components decreased INN retention. INN/meter2 of lung, OX, and AF at 3 hr were 0.26 +/- 0.07%, 0.06 +/- 0.02%, and 6.17 +/- 3.94%, and significantly lower on a heparin coated filter (2.14 +/- 1.30%). Capillary surface areas of viscera and connective tissues (lung, 100; liver, 134; spleen, 20; heart, 7; skeletal muscle, 92; fat, 12; bone, 3; bone marrow, 5; brain, 0.1 meter2) were estimated from distribution of activated INN in pigs. Lung INN retention was much higher than that of the polymer surfaces of OX/AF, indicating the role of cell adhesion molecules on INN retention on endothelial cells of lung and viscera. By direct continuous monitoring and quantitation of INN at the end of CPB, a sensitive technique for quantitation of neutrophil kinetics, margination, and retention during CPB was developed.

Rate constants of embolization and quantitation of emboli from the hollow-fiber oxygenator and arterial filter during cardiopulmonary bypass

A direct technique was developed to estimate parameters related to half life (T1/2) and rate constants of embolization (RCE) and to quantitate emboli of three sizes (small, medium, and large) shed from the oxygenator and arterial filter during cardiopulmonary bypass (CPB). Cardiopulmonary bypass was performed in 16 Yorkshire pigs divided as follows: systemic heparin group (SHG:6), systemic heparin/heparinized circuit group (SH/HCG:5), and Iloprost (Bentley) (2 ng/kg/min)/heparinized circuit group (IHCG:5) with In-111 labeled autologous platelets. The anesthetized pigs (20-25 kg) underwent CPB at 2.5-3.0 L/min for 3 hours. Pigs were injected with In-111 platelets (300-420 microCi) 24 hours before CPB. Cardiopulmonary bypass was instituted with a roller pump and hollow-fiber oxygenator, and thrombosis and embolization on the oxygenator and filter were monitored by a calibrated Geiger probe (WMB Johnson Associates, Montvale, NJ). The radioactivity in the oxygenator and filter reached peak values at 25-45 min after CPB; the radioactivity then declined in the oxygenator but remained at a steady state in the filter, suggesting continuous embolization at the same rate of trapping. The curve stripping technique of the normalized radioactivity time curve of the oxygenator was used for RCE estimation of different sized emboli shed from the oxygenator; 42% of thrombus embolized from the oxygenator in SHG with three rate constants, with T1/2s of 12 min, 42 min, and 13 hr; the SH/HCG embolized 35% with T1/2 of 78 min, and the IHCG embolized 30%, with a T1/2 of 22 min. This indicates that there is less embolization in the IHCG.(ABSTRACT TRUNCATED AT 250 WORDS)

Reduction of neutrophil margination by L-arginine during hypothermic cardiopulmonary bypass in a pig model.

Nitric oxide generation by L-arginine (2 mg/kg/min) infusion during cardiopulmonary bypass (CPB) increases blood flow to all organs and reduces cytokine induced organ damage by reducing the level of marginating neutrophils (Ns). The N-trapping in the oxygenator (OX), arterial filter (AF), cardiotomy reservoir (CR), and N-margination were quantified with indium 111 labeled autologous neutrophils (INN) in nine groups of 40 Yorkshire pigs (30-35 kg). Cardiopulmonary bypass (180 min or 90 min CPB, 90 min reperfusion) was carried out at 2.5-3.5 L/min and at two temperatures (18 degrees C, 28 degrees C). The INN (650-780 microCi) was administered intravenously 15 mins before CPB. All pigs received heparin systemically (activated coagulation time > 400 secs); CPB was instituted with a roller pump, OX (Univox 1.8 m2), AF (0.25 m2), and CR (BCR-3500, Bentley Lab, Irvine, CA). The INN distribution in the device (OX, AF, CR) and organs was imaged with a gamma camera and measured with an ion chamber and a gamma counter. The LA infusion decreased N-trapping, estimated as the percent of injected INN (mean +/- standard deviation), in OX from control (2.7 +/- 2.02)% to (0.94 +/- 0.29)%, and margination in lung from control (48 +/- 4)% to minimal levels (23 +/- 2)% (p < 0.01). In the CPB reperfusion group, a beneficial effect was observed at LA low dose and toxicity of higher N-margination at 15 mg/ kg/min. Neither CPB temperature nor Leumedin affected N-margination significantly.

Quantitation of platelet loss with indium-111 labeled platelets in a hollow-fiber membrane oxygenator and arterial filter during extracorporeal circulation in a pig model

Platelet consumption in a hollow-fiber membrane oxygenator (HFMO) and arterial filter (AF) during cardiopulmonary bypass (CPB) was quantified in five pigs using Indium-111 labeled autologous platelets. Platelet labeling was performed 20-24 hours before CPB. After general endotracheal anesthesia, the pigs were systemically heparinized and were placed on CPB via a median sternotomy. After 3 hours of CPB, radioactivity was quantified with a gamma camera and an ionization chamber. The percent of injected dose (mean +/- SD) was 0.79 +/- 0.45 in the HFMO, 2.52 +/- 0.93 in AF, 4.3 +/- 1.2 in blood loss during CPB. Platelet consumption in HFMO during CPB was lower than in bubble oxygenators (19%) or silicone membrane oxygenators (12%) as observed in previous studies.

Quantification of Device Adherent, Circulating, and Organ Pool of Thrombin and Fibrinogen After Cardiopulmonary Bypass in a Pig Model

The pool of thrombin and fibrinogen in circulation, in organs, and on cardiopulmonary bypass devices was quantified during and after cardiopulmonary bypass in four groups of 24 Yorkshire pigs (weight, 30-35 kg); two groups of 10 unoperated pigs were used as controls. Thrombin-alpha and fibrinogen were iodinated with 125iodide using an iodogen transfer technique; 250-300 microCi of these tracers were injected intravenously 1 hr before cardiopulmonary bypass. All pigs were systematically heparinized (activated clotting time > 400 sec); cardiopulmonary bypass was performed at 2.5-3.5 L/min at 28 degrees C using a centrifugal pump, oxygenator (Bentley Univox 1.8 m2; Bentley Inc., Irvine, CA), arterial filter (0.25 m2), and cardiotomy reservoir (BMR 3500) for 90 min, followed by a 90 min reperfusion and 180 min of cardiopulmonary bypass. Iodinated thrombin-alpha and fibrinogen in intact organs and samples of blood, organs, tissues, and oxygenator-arterial filter-cardiotomy reservoir were quantified with an ion chamber and a gamma counter, respectively. The percent of injected iodinated thrombin-alpha and fibrinogen dose (mean +/- SD) in organs and cardiopulmonary bypass devices of all groups of cardiopulmonary bypass pigs was calculated. Thrombin generated at the small area of surgical wounds (0.016-0.038 m2), and fibrin deposited on surfaces of cardiopulmonary bypass devices (2.59 m2), initiate and propagate thrombus formation and embolization. The protein level reached saturation values on all cardiopulmonary bypass devices at 180 min. High levels of thrombin and fibrinogen-fibrin circulate in blood and organs, and are adsorbed on cardiopulmonary bypass devices; this large blood pool of pro-coagulants in the cardiac cradle, tissues, and perfused organs may account for thrombi and emboli during and after cardiopulmonary bypass.

Effect of heparin reversal and fresh platelet transfusion on platelet emboli post-cardiopulmonary bypass in a pig model.

Heparin reversal by protamine and fresh platelet transfusion may decrease bleeding complications post-cardiopulmonary bypass (CPB) and may increase the level of organ trapped platelet emboli. Platelet emboli were quantified in two groups of 12 Yorkshire pigs (30-35 kg), where 111indium labeled autologous platelets (INPLT: 850-1,200 microCi) were injected intravenously before and after CPB (BCPB, ACPB), and the platelet emboli level in intact organs and their samples (brain, heart, kidneys, lung, liver, and spleen) was quantified with an ion chamber and a gamma counter, respectively. All pigs were systemically heparinized (ACT > 400 sec). CPB was carried out at 2.5-3.5 L/min at 28 degrees C using a centrifugal pump, an oxygenator (OX:Bentley Univox 1.8 m2), an arterial filter (AF:0.25 m2), and a cardiotomy reservoir (CR: BMR 250) for 90 min. Heparin was reversed with an equivalent dose of protamine. The percent of INPLT dose (ID%, mean +/- SD) in organs of BCPB and ACPB pigs was calculated. The sequence of platelet emboli on a unit weight basis (ID%/g) had the following order: Spleen > Liver > Lung > Kidneys > Heart > Brain. The presence of significantly higher levels of emboli in brain, heart, and kidneys in the ACPB than the BCPB group suggest that platelet transfusion after heparin reversal with protamine may increase the risk of platelet emboli. However, it is an acceptable risk for patients having bleeding complications post-CPB.